Evolutionary theory proposes that mitochondria and plastids originated by engulfment or cell fusion of prokaryotes by eukaryotes. As this relationship evolved, the size of the bacterial DNA genome decreased and the functions of genes lost from the bacterial genome were assumed by the eukaryotic chromosome (Cavalier-Smith. (1987) Ann. NY Acad. Sci. 503:55-71). Support for this theory is found in the fungus, Geosiphon pyriforme, which contains in its hyphal system cyanobacteria belonging to the genus, Nostoc, but which retain the capacity for autonomous growth and replication (Mollenhauer. (1992) Geosiphon pyriforme, In Algae and Symbiosis. Biopress Ltd., Bistol, pp. 339-351). Additional support is found in algae which have plastids containing DNA that has a significant level of homology and similar gene organization to cyanobacteria but the plastids have lost most of the cyanobacterial genes to the cell nucleus (Douglas. (1994) Chloropast Origins and Evolution, In Molecular Biology of Cyanobacteria, vol. 1 (Bryant, ed.) Kluwer Academic Publishers, Boston, pp. 91-118). The reason and mechanism for the relocation of a large proportion (greater than 90%) of the bacterial genes to the nucleus are unknown (Valentin et al. (1992) Phylogenetic origin of the plastids, In Origins of Plastids (Lewin, ed.) Chapman and Hall, New York, pp. 193-221).
It would therefore be useful to develop a system to introduce an entire prokaryotic genome into a eukaryotic organism and to study the interactions of the two genomes and the effect this has on both organisms. Preferably, such a system would permit both nuclear and extra-nuclear localization of the bacterial genome. This system also would provide a model for the evolution of mitochondria, chloroplasts, and other plastids.
The present invention presents new technology that can be used to transfer entire bacterial chromosomes into yeast or other eukaryotic organisms in such a manner that they become functional linear artificial chromosomes and, furthermore, may become compartmentalized bacteria or organelle-like structures. The bacterial chromosome will be expressed partially in the nucleus and in the bacterial organelle and provides new and useful pathways to the eukaryote host immediately after formation of the hybrid cell or after selections for specific desired functions normally done by the prokaryote alone as well as new functions. These new vectors and methods additionally provide a means to efficiently introduce very large segments of DNA into eukaryotic cells without extracellular manipulation.